P998: TRETINOIN AND CALCITRIOL ENHANCE TREATMENT REGIMENS FOR JUVENILE MYELOMONOCYTIC LEUKEMIA IN EX VIVO DRUG SENSITIVITY ASSAYS
Authors: Elliot Stieglitz, Christine Gu, Anukriti Dhar, Juwita Huebner, Kamran Ali, Markus Lacher
Journal: HemaSphere (2023)

Topic: 15. Myeloproliferative neoplasms - Biology & Translational Research Background: Juvenile myelomonocytic leukemia (JMML) is a rare and aggressive pediatric malignancy with features of both myelodysplasia and myeloproliferation. In nearly all cases, a RAS pathway gene (PTPN11, NRAS, KRAS, NF1, or CBL) is mutated. For most patients, the only curative treatment option is hematopoietic stem cell transplantation (HSCT). As pre-HSCT treatments, fludarabine (FLU) plus cytarabine (ARA-C) and 5-azacitidine (5-AZA) monotherapy are commonly used but are not universally effective. There is a need to identify additional, more effective pre-HSCT regimens in JMML. Aims: We aimed at identifying novel, anti-leukemic drug combinations in JMML by employing a high-throughput, flow cytometry-based predictive precision medicine platform (PPMP). Methods: We screened 130 drug combinations (DCs) (dual and triple) on peripheral blood or bone marrow samples from 29 JMML patients to identify combinations that significantly reduced the vehicle-normalized blast (NormBlast) counts relative to their individual components (ICs). For samples demonstrating this effect (referred to as “DC-advantage”) and belonging to such combinations, effect sizes were calculated to quantify the combinatorial effects of the combinations, whereby effect size was defined as the NormBlast difference between the DC and its IC with the lowest NormBlast count. Informed consent was obtained for all patients. Results: We identified 37 drug combinations with combinatorial effects. Tretinoin (TRET, all-trans retinoic acid), a differentiation agent, enhanced the anti-leukemic effects of FLU + ARA-C in 91% (21/23) of patients and of 5-AZA in 78% (21/27) of patients. Similarly, calcitriol (CALC), another differentiating agent, enhanced the anti-leukemic effect of 5-AZA in 85% (23/27) of patients. For patients with a DC-advantage, the median NormBlast values were reduced as follows: for FLU + ARA-C by 88%, and with the inclusion of TRET by 91% (median effect size: 3%); for 5-AZA by 27%, and with the inclusion of TRET by 51% (median effect size: 15%), or with the inclusion of CALC, by 46% (median effect size: 13%). For the patients with both a DC-advantage and NormBlast counts >30% following ex vivo treatment with FLU + ARA-C or 5-AZA, addition of TRET reduced the median NormBlast values from 32% to 28% (n=3), or from 73% to 49% (n=21), respectively. The addition of CALC to 5-AZA also reduced the NormBlast value, from 73% to 54% (n=23). Figure 1 outlines the respective NormBlast values and effect sizes following ex vivo treatment with the indicated conditions. Addition of the differentiating agent (TRET or CALC) to 5-AZA resulted in a larger blast-reduction effect compared to FLU + ARA-C. However, the reduction of the NormBlast count by FLU + ARA-C without the differentiating agent was greater than by 5-AZA alone. Summary/Conclusion: We have shown the utility of a high-throughput PPMP for identifying drug combinations potentially more effective than currently used JMML pre-HSCT regimens. Using the largest collection of primary JMML samples tested ex vivo in the literature, we highlight that the addition of tretinoin to FLU + ARA-C or to 5-AZA, or calcitriol to 5-AZA, further reduces the ex vivo blast counts compared to treatment with only FLU + ARA-C, or 5-AZA, respectively.Keywords: JMML, Ex vivo, ALL-trans retinoic acid (ATRA), Drug sensitivity